DK2650364T3 - Hafniaphytasevarianter - Google Patents

Hafniaphytasevarianter Download PDF

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Publication number
DK2650364T3
DK2650364T3 DK13170321.7T DK13170321T DK2650364T3 DK 2650364 T3 DK2650364 T3 DK 2650364T3 DK 13170321 T DK13170321 T DK 13170321T DK 2650364 T3 DK2650364 T3 DK 2650364T3
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Denmark
Prior art keywords
phytase
variant
seq
activity
leu
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DK13170321.7T
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English (en)
Inventor
Soeren Flensted Lassen
Lars Kobberoee Skov
Maria Leonardo De
Tomoko Matsui
Allan Noergaard
Jesper Vind
Esben Peter Friis
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Novozymes As
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    • CCHEMISTRY; METALLURGY
    • C12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12NMICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/14Hydrolases (3)
    • C12N9/16Hydrolases (3) acting on ester bonds (3.1)
    • AHUMAN NECESSITIES
    • A23FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23KFODDER
    • A23K20/00Accessory food factors for animal feeding-stuffs
    • A23K20/10Organic substances
    • A23K20/189Enzymes
    • AHUMAN NECESSITIES
    • A23FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23KFODDER
    • A23K50/00Feeding-stuffs specially adapted for particular animals
    • A23K50/30Feeding-stuffs specially adapted for particular animals for swines
    • AHUMAN NECESSITIES
    • A23FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23KFODDER
    • A23K50/00Feeding-stuffs specially adapted for particular animals
    • A23K50/70Feeding-stuffs specially adapted for particular animals for birds
    • A23K50/75Feeding-stuffs specially adapted for particular animals for birds for poultry
    • YGENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02EREDUCTION OF GREENHOUSE GAS [GHG] EMISSIONS, RELATED TO ENERGY GENERATION, TRANSMISSION OR DISTRIBUTION
    • Y02E50/00Technologies for the production of fuel of non-fossil origin
    • Y02E50/10Biofuels, e.g. bio-diesel

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  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Polymers & Plastics (AREA)
  • Engineering & Computer Science (AREA)
  • Zoology (AREA)
  • Animal Husbandry (AREA)
  • Food Science & Technology (AREA)
  • Health & Medical Sciences (AREA)
  • Birds (AREA)
  • Organic Chemistry (AREA)
  • Genetics & Genomics (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Wood Science & Technology (AREA)
  • Microbiology (AREA)
  • General Engineering & Computer Science (AREA)
  • General Health & Medical Sciences (AREA)
  • Biochemistry (AREA)
  • Molecular Biology (AREA)
  • Biotechnology (AREA)
  • Medicinal Chemistry (AREA)
  • Biomedical Technology (AREA)
  • Enzymes And Modification Thereof (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)
  • Fodder In General (AREA)

Claims (20)

1. Fremgangsmåde til fremstilling af en phytasevariant af en reference- eller udgangsphytase med forbedrede egenskaber med hensyn til pH-profil og med mindst 90% identitet med SEQ ID NO:2, hvor varianten har mindst en substitution, insertion eller deletion i en eller flere af positionerne 92, 48, 26, 45, 49, 68, 100, 162, 217, 228, 304, 308 og 358, hvor positionerne svarer til positionerne i phytasen med aminosyrerne 1 -413 i SEQ ID NO:2, hvorved fremgangsmåden omfatter a) mutation af DNA’et eller genet, der koder for udgangsphytasen på en måde, så DNA’et eller genet koder for substitutionen, insertionen og/eller deletionen, b) operabel forbindelse af DNA’et eller genet til en eller flere kontrolsekvenser, der styrer fremstillingen af phytasen i en passende ekspressionsvært til fremstilling af en DNA-konstruktion eller en rekombinant ekspressionsvektor, c) overføring af konstruktionen eller vektoren til en passende vært, d) kultivering af værten til fremstilling af phytasevarianten, og e) udvinding af phytasen.
2. Fremgangsmåde ifølge krav 1, hvor modifikationerne er valgt fra følgende substitutioner eller insertioner: 92Y, 48H,W,N, 26R,Q,H, 49L, 100W, 162N,R, 217S,G, 304V, 308A og 358C.
3. Phytasevariant med mindst 90% identitet med SEQ ID NO:2 og med forbedrede egenskaber med hensyn til pH-profil, hvor pH-profilen er bestemt ved 37°C, ved måling af aktiviteten i forhold til den maksimale aktivitet, er forbedret sammenlignet med pH-profilen for phytasen fra SEQ ID NO:2, og fremstillet i henhold til metoden ifølge krav 1 eller 2, med det forbehold at phytasen ikke er Hafnia alvei phytasen med aminosyrerne 1-413 fra SEQ ID NO:2.
4. Phytase ifølge krav 3 omfattende mindst en modifikation valgt fra følgende: 228C, 325C, 358C og 363C.
5. Phytase ifølge krav 4 omfattende mindst et sæt af modifikationer valgt fra følgende: 325C/358C og 228C/363C.
6. Phytase ifølge krav 4 eller 5 omfattende mindst en yderligere modifikation i en position valgt fra følgende: 1,4, 5, 6, 7, 9, 10, 12, 16, 18, 25, 26, 27, 28, 29, 30, 31,32, 37, 38, 39, 40, 41,45, 48, 49, 55, 59, 64, 68, 69, 70, 71,72, 74, 75, 76, 77, 78, 79, 80, 81,82, 83, 89, 91,92, 93, 95, 96, 97, 98, 100, 103, 108, 109, 110, 111, 112, 113, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 128, 130, 131, 132, 133, 134, 136, 137 ,138, 140, 144, 145, 146, 147, 148, 149, 151, 152, 153, 154, 155, 156, 158, 159, 160, 161, 162, 163, 168, 173, 175, 180, 181, 182, 183, 184, 185, 186, 187, 189, 190, 192, 193, 194, 195, 196, 198, 199, 200, 202, 203, 204, 205, 206, 207, 208, 209, 211,215, 217, 219, 221,227, 230, 233, 234, 235, 238, 239, 240, 241,242, 243, 244, 245, 246, 247, 248, 249, 251,256, 258, 260, 261,266, 268, 270, 279, 284, 285, 286, 287, 288, 289, 290, 292, 293, 294, 295, 296, 297, 298, 299, 301,303, 304, 308, 310, 312, 313, 314, 316, 318, 319, 320, 322, 324, 335, 344, 345, 346, 347, 348, 354, 355, 356, 360, 362, 364, 365, 366, 367, 369, 371,372, 373, 375, 376, 378, 382, 383, 384, 385, 386, 387, 388, 389, 390, 391,394, 395, 396, 397, 400, 401,403, 404, 406, 408, 409, 411,412 og 413.
7. Phytase ifølge et hvilket som helst af kravene 3-6 omfattende mindst en modifikation valgt fra følgende: 49L.
8. Phytase ifølge krav 3-7 omfattende mindst en modifikation valgt fra følgende: T35C, P36C, F63C, L172C, S177C, P178C, F224C, H228C, A236C, Q326C, P331C, T358C, L368C og A374C.
9. Phytase ifølge krav 8 omfattende mindst et sæt af modifikationer valgt fra følgende: F8C/D343C, P178C/D33C, F63C/L368C, F224C/A236C, P331C/Q326C, T358C/G325C, H228C/H363C og L368C/A374C.
10. Phytase ifølge et hvilket som helst af kravene 6-9 omfattende mindst en modifikation valgt fra følgende: I49L.
11. Phytase ifølge et hvilket som helst af kravene 6-10 omfattende mindst en modifikation valgt fra: Q162N og A304V.
12. Isoleret nukleinsyresekvens omfattende en nukleinsyresekvens, der koder for phytasen ifølge et hvilket som helst af kravene 3-11.
13. Nukleinsyrekonstruktion omfattende nukleinsyresekvensen ifølge krav 12 operabelt forbundet med en eller flere kontrolsekvenser, der styrer fremstillingen af phytasen i en passende ekspressionsvært.
14. Rekombinant ekspressionsvektor omfattende nukleinsyrekonstruktionen ifølge krav 13.
15. Rekombinant værtscelle omfattende nukleinsyrekonstruktionen ifølge krav 13 og/eller ekspressionsvektoren ifølge krav 14.
16. Sammensætning omfattende mindst en phytase ifølge et hvilket som helst af kravene 3-11, og (a) mindst et fedtopløseligt vitamin; (b) mindst et vandopløseligt vitamin; og/eller (c) mindst et spormineral.
17. Sammensætning ifølge krav 16 yderligere omfattende mindst et enzym valgt fra følgende gruppe af enzymer: amylase, phytase, phosphatase, xylanase, galactanase, alpha-galactosidase, protease, phospholipase, og/eller beta-glucanase.
18. Sammensætning ifølge et hvilket som helst af kravene 16-17, hvilken sammensætning er et dyrefoderadditiv.
19. Fremgangsmåde til fremstilling af et fermenteringsprodukt omfattende (a) fermentering af et materiale indeholdende kulhydrat under tilstedeværelse af en phytase ifølge et hvilket som helst af kravene 3-11 og ved anvendelse af en fermenteringsmikroorganisme og (b) fremstilling af fermenteringsproduktet eller fermenteringsbiproduktet fra materialet indeholdende det fermenterede kulhydrat.
20. Fremgangsmåde ifølge krav 19, hvor fermenteringsproduktet er ethanol, øl, vin eller tørret bærme (distillers dried grains (DDG)).
DK13170321.7T 2008-09-26 2009-09-28 Hafniaphytasevarianter DK2650364T3 (da)

Applications Claiming Priority (2)

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EP08165245 2008-09-26
EP09783496.4A EP2342323B1 (en) 2008-09-26 2009-09-28 Hafnia phytase variants

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DK2650364T3 true DK2650364T3 (da) 2015-06-08

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DK09783496.4T DK2342323T3 (da) 2008-09-26 2009-09-28 Hafnia phytasevarianter
DK15166896.9T DK2933329T3 (da) 2008-09-26 2009-09-28 Hafniaphytasevarianter

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US (2) US8206962B2 (da)
EP (3) EP2650364B1 (da)
CN (1) CN102165059A (da)
AU (1) AU2009295852B2 (da)
BR (1) BRPI0919314A2 (da)
CA (1) CA2736321C (da)
DK (3) DK2650364T3 (da)
ES (3) ES2426717T3 (da)
MX (1) MX2011003132A (da)
PL (1) PL2342323T3 (da)
WO (1) WO2010034835A2 (da)

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ES2636369T3 (es) 2017-10-05
PL2342323T3 (pl) 2013-11-29
US8507240B2 (en) 2013-08-13
MX2011003132A (es) 2011-04-21
EP2650364B1 (en) 2015-05-20
EP2342323A2 (en) 2011-07-13
BRPI0919314A2 (pt) 2015-08-11
ES2544645T3 (es) 2015-09-02
US8206962B2 (en) 2012-06-26
CN102165059A (zh) 2011-08-24
CA2736321C (en) 2018-09-11
WO2010034835A2 (en) 2010-04-01
WO2010034835A3 (en) 2010-06-24
CA2736321A1 (en) 2010-04-01
EP2933329A1 (en) 2015-10-21
DK2933329T3 (da) 2017-07-24
ES2426717T3 (es) 2013-10-24
EP2342323B1 (en) 2013-06-05
AU2009295852A1 (en) 2010-04-01
EP2933329B1 (en) 2017-05-17
DK2342323T3 (da) 2013-09-08
AU2009295852B2 (en) 2015-06-25
US20100083392A1 (en) 2010-04-01
EP2650364A1 (en) 2013-10-16
US20120225468A1 (en) 2012-09-06

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